Development of Chimeric Molecules That Degrade the Estrogen Receptor

Development of chimeric molecules that degrade the estrogen receptor using decoy oligonucleotide ligands Miyako Naganuma,†,‡,§ Nobumichi Ohoka,||,* Genichiro Tsuji,† Haruna Tsujimura,†,‡ Kenji Matsuno,§ Ta- kao Inoue,|| Mikihiko Naito,** and Yosuke Demizu†,‡,*

†Division of Organic Chemistry, National Institute of Health Sciences, Kanagawa, Japan ‡Graduate School of Medical Life Science, Yokohama City University, Kanagawa, Japan §Department of Chemistry and Life Science, Kogakuin University, Tokyo, Japan ||Division of Molecular Target and Gene Therapy Products, National Institute of Health Sciences, Kanagawa, Japan **Laboratory of Targeted Protein Degradation, Graduate School of Pharmaceutical Sciences, The University of Tokyo, Tokyo, Japan KEYWORDS: ubiquitin-proteasome system; protein knockdown; decoy; transcription factors; estrogen receptor

ABSTRACT: Targeted protein degradation using chimeric small molecules, such as proteolysis-targeting chimeras (PROTACs) and specific and nongenetic inhibitors of apoptosis protein [IAP]-dependent protein erasers (SNIPERs), has attracted attention as a method to degrade intracellular target proteins via the ubiquitin-proteasome system (UPS). These chimeric molecules target a variety of proteins using small molecules that can bind to the proteins. However, it is difficult to develop such degraders in the absence of suitable small molecule ligands for the target proteins, such as for transcription factors (TFs). Therefore, we constructed the chimeric molecule LCL-ER(dec), which consists of a decoy oligonucleotide that can bind to the estrogen receptor a (ERa) and an IAP ligand, LCL161 (LCL), in a click reaction. LCL-ER(dec) was found to selectively degrade ERa via the UPS. These findings will be applicable to the development of other oligonucleotide-type degraders that target different TFs.

The identity of many target proteins that are involved in the pathogenesis of various diseases has been elucidated, and Ubiquitination E3 LCL161, VH032, various drug discovery techniques have been developed to tar- ligase Ligand 1, 2 Pomalidomide get these proteins. In particular, the use of targeted protein Ligand Ub O Decoy of ERa O Ub degradation has recently been attracting attention. This technol- Ub O Ub Ubiquitin ogy uses chimeric molecules called proteolysis-targeting chi- Ub N N ERa Proteasome meras (PROTACs) or specific and nongenetic inhibitors of N Degradation O apoptosis protein [IAP]-dependent protein erasers (SNIPERs) O P O OH to degrade target proteins in cells via the ubiquitin-proteasome Figure 1. Decoy-type PROTACs degrade the ERa via system (UPS).3, 4 These chimeric molecules consist of a ligand the UPS. for the target protein (protein of interest, POI) and an E3 ligand. The proximity of the POI and E3 ligase acts as a molecular me- diator, thereby promoting the ubiquitination and subsequent transcriptional factors (TFs), such as the estrogen receptor a degradation of the POI.2 Small molecules, such as LCL161 (ERa)5 and neurogenic locus notch homolog protein 1 (LCL) for IAPs, VH032 (VH) for von Hippel-Lindau protein (NOTCH1),6 by introducing an oligopeptide that interacts with (VHL), and pomalidomide (POM) for cereblon (CRBN) are of- the surface of the POI. This peptide-based protein knockdown ten used as ligands for E3 ligases.3 A small molecule that is strategy is also applicable to several types of POIs that have no known to bind to a POI can be used to develop chimeric mole- specific small-molecule ligands. cules, and many chimeric degraders using small molecules as Other molecules that can bind to protein surfaces include 3 POI ligands have been developed to date. Because the only ac- oligonucleotides, such as aptamers7 and decoys8. If such oligo- tivity required for the ligand is for it to be able to bind to the nucleotides can be used as POI ligands in the development of POI, this technology can target proteins that have no enzymatic chimeric degraders, the number of target proteins that can be activity (i.e., for which inhibitors cannot be developed). How- degraded would be expanded. It is particularly advantageous to ever, it is difficult to develop small molecule-based degraders use a decoy as the ligand when targeting TFs, because the decoy against a POI in the absence of appropriate ligands. An alterna- can use the known DNA-binding sequence of the target TF, tive approach is to use molecules, such as peptides, that can bind making it easy to design the ligand. In the present study, we to the protein surfaces as POI ligands. Several groups have re- developed decoy-based chimeric degraders that target the ERa cently reported peptide-based degraders that target

Figure 2. Molecules synthesized in this study. as a model TF (Figure 1). The ERa is a member of the nuclear with an AGCT repeat sequence. The chimeric molecules LCL- hormone receptor superfamily9 and can bind directly to DNA to Scrbl-1, LCL-Scrbl-2, and LCL-Scrbl-3 were synthesized in act as a TF.10 In addition, ERa homodimers can form transcrip- a similar manner as LCL-ER(dec). A non-degradable control tional complexes on the DNA response sequence and regulate NMeLCL-ER(dec) containing an N-methylated analog of the expression of target genes by recruiting co-regulators.10 We LCL161, and the fluorescein (FAM)-labeled decoy FAM- have previously developed small molecule-based chimeric de- ER(dec) for use in a competitive fluorescence polarization as- graders against ERa11, 12 using the ERa antagonist 4-hydroxyta- say were also synthesized (Figure 2). The detailed synthetic moxifen (4-OHT), and peptide-based ERa degraders5 using a protocols of these molecules are described in the supporting in- coactivator motif that interacts with the ERa surface. Therefore, formation. we chose ERa, as a model TF, as the target for the development The preferred higher-order structures of the synthesized of oligonucleotide-type chimeric degraders. Specifically, we ER(dec) and the chimeric molecules LCL-ER(dec), VH- designed a thermodynamically stable double-stranded decoy, ER(dec), and POM-ER(dec) were analyzed using CD spectra. ER(dec), based on the sequence of the estrogen-responsive el- Each decoy showed a negative maximum at approximately 240 ement (ERE)13 that is known to bind strongly to the ERa. This nm and a positive maximum at approximately 280 nm, indicat- decoy was conjugated with three different E3 ligase ligands, ing that LCL-ER(dec), VH-ER(dec), and POM-ER(dec) LCL, VH, and POM, to construct the degraders LCL-ER(dec), formed typical right-handed B-type structures and the terminal VH-ER(dec), and POM-ER(dec), respectively. The structural modification of ER(dec) did not affect the higher-order struc- properties, binding affinities, and ERa-degradation activities of ture (Figure S4). The melting temperature (Tm) values of these degraders were evaluated. ER(dec), LCL-ER(dec), VH-ER(dec), and POM-ER(dec) were 64.5, 64.3, 66.9, and 62.9 °C, respectively, which indi- The ERa-binding decoy ER(dec) (21 residues) was de- cated that these molecules had similar conformational stability signed using information from the DNA sequence and co-crys- to ER(dec) (Figure S6). tal X-ray structure of the ERa,14 and an ERE from the Xenopus vitellogenin A2 gene.13 The 5'-end of the sense strand 5'- The binding affinities of ER(dec), LCL-ER(dec), VH- GTCAGGTCACAGTGACCTGAT-3' of ER(dec) was modi- ER(dec), and POM-ER(dec) to the ERa were evaluated by a fied with a hexynyl group. To construct the E3 ligase ligands, a competitive fluorescence polarization assay. For the evaluation, PEG3 linker with an azide group was attached to LCL, VH15, the compounds were added to a buffer system containing ERa and POM16. Then, the above sense strand with the alkyne was and FAM-ER(dec), and the inhibitory concentration was cal- conjugated to each azide-containing E3 ligase ligand by a cop- culated as the IC50 value. The IC50 values were 83.2 nM for per-catalyzed click reaction. Subsequent hybridization with the ER(dec), 37.9 nM for LCL-ER(dec), 39.3 nM for VH- antisense strand 5'-ATCAGGTCACTGTGACCTGAC-3' af- forded the desired chimeric molecules LCL-ER(dec), VH- ER(dec), and POM-ER(dec). In addition, three ERa non-binding decoys were designed with the same base composition as ER(dec): Scrbl-1 with an AT repeat and GC repeat sequence; Scrbl-2 with a completely randomized sequence; and Scrbl-3

Figure 3. Degradation of ERa via UPS by the synthesized decoys. (A, B) The synthesized decoys selectively induce reduction of ERa protein level. MCF-7 cells were transfected with the indicated concentrations of LCL-ER(dec), VH-ER(dec), or POMFigure-ER(dec) 3. Degradation for 24 h. (C) of The the EReffecta via of the UPS UPS inhibitors by the synthesized on the LCL decoys.-ER(dec) (A,- inducedB) The synthesized reduction of decoys ERa. selectivelyMCF-7 cells induce wered transfecteda reduction within the the ER indicateda protein concentrationslevel. MCF-7 cells of LCL were-ER(dec) transiently in transfectedthe presence with or absence the indicated of 10 concentrationsµM MG132 or ofMLN7243 LCL-ER(dec) for 24, h.VH (D-ER(dec)-F) Degradation, or POM of- ER(dec)ERa protein for by 24 LCL h. (C)-ER(dec) The effect is IAP of ligan UPSd -dependentinhibitors onand theDNA LCL sequence-ER(dec)-dependent.-induced MCF reduction-7 cells of wereERa levelstransfected. MCF with-7 cells the wereindicated transiently concentrations transfected of withLCL the-ER(dec) indicated, ER(dec) concentrations, NMeLCL of LCL-ER(dec)-ER(dec), LCL in- Scramblethe presence-1, LCLor ab-- Scramblesence of 10-2 µ, Mor MG132LCL-Scramble or MLN7243-3 for for24 24h. Wholeh. (D–F)-cell Degradation lysates were of analyzedthe ERa proteinby Western by LCL blotting-ER(dec) with isthe IAP indicated-ligand antibodies; and DNA- representativesequence dependent. data are MCF shown.-7 cells Numbers were belowtransiently the ER transfeca panelsted represent with the ER/actinindicated ratios, concentrations normalized of by LCL designating-ER(dec) the, ER(dec) expres-, sionNMeLCL from -theER(dec) vehicle, LCL control-Scramble condition-1, (conditionsLCL-Scramble without-2, ordecoys) LCL- Scrambleas 100%. -The3 for changes 24 h. Whole in protein-cell lysateslevels were were reproducibleanalyzed by betweenwestern blottingthe two independentwith the indicated experiments. antibodies; Data representative in the bar graph data are are the shown. mean ofTh twoe numbers results. below the ERa panels represent the ER/actin ratios, normalized by designating the expression using the vehicle control (condition without a decoy) as 100%. The changes in protein levels were reproducible between two independent experiments. The data in the bar graph are the mean of two results. Abbreviations; AR: androgen receptor, AhR: aryl hydrocarbon receptor, BRD4: bromodomain-containing protein 4, GAPDH: glyceraldehyde 3-phosphate dehydrogenase, CRABP2: cellular retinoic acid binding protein 2.

ER(dec), and 50.5 nM for POM-ER(dec). These results indi- 6 LCL-Scrbl-2 3.28×103 cated that the binding activity of the decoy ER(dec) to the ERa 7 LCL-Scrbl-3 8.16×103 was not weakened by the linking of ER(dec) to the E3 ligase ligand, and the compounds had sufficient binding ability to the ERa regardless of the type of the E3 ligase ligand (Table 1). To investigate whether LCL-ER(dec), VH-ER(dec), and The binding activities of LCL-Scrbl-1–3 to the ERa were con- POM-ER(dec) have degradation activity against the ERa, the siderably weaker than that of ER(dec) composed of the consen- effect of these E3 ligand-decoy conjugates on the protein level sus sequence, but some non-specific binding was observed at of ERa was evaluated by western blotting analysis using MCF- high concentrations (Figures S8 and S9). 7 breast cancer cells. All the decoys reduced the ERa protein level after 24 h of transfection, and LCL-ER(dec) had the most effect compared with the other decoys (Figure 3A). To investi- Table 1. ERa binding affinity (IC50) of ER(dec), LCL- gate the selectivity for the target protein, the effect of these de- ER(dec), VH-ER(dec), POM-ER(dec), LCL-Scrbl-1, LCL- coys on the levels of different proteins was examined (Figure Scrbl-2, and LCL-Scrbl-3. 3B). None of the decoys effectively reduced the levels of other Entry Compound IC50 (nM) transcription factors (AR, AhR, and NF-kB p65), transcription- 1 ER(dec) 67.0 related factors (p300 and BRD4), or proteins not related to transcription (CRABP2, GAPDH, and b-actin), indicating that 2 LCL-ER(dec) 37.9 these decoys selectively reduced the protein level of ERa. 3 VH-ER(dec) 50.4 We next investigated the mechanism of the ERa reduction 4 POM-ER(dec) 39.3 by focusing on LCL-ER(dec), which showed the highest activ- 5 LCL-Scrbl-1 10.8×103 ity, as shown in Figure 3A. The LCL-ER(dec)-induced reduction of the ERa was abrogated by co-treatment with the

In summary, we synthesized LCL-ER(dec) with the aim of developing a chimeric degrader using a decoy ligand that can be applied to a variety of TFs. For the target protein, we selected the ERa, which has previously been shown to be degraded by chimeric molecules containing small molecules or peptides. The three chimeric molecules LCL-ER(dec), VH-ER(dec), and POM-ER(dec) were synthesized by conjugating the ERa decoy ER(dec) with LCL161, VH032, and POM, E3 ligase ligands commonly used in PROTACs and SNIPERs. Among these three chimeric molecules, LCL-ER(dec) showed the highest ERa degradation-inducing activity as assessed by western blotting. In contrast, the ERa-non-binding chimeric molecules, LCL-Scrbl-1–3, did not reduce the protein levels of ERa, suggesting that LCL-ER(dec) binds to the ERa in a sequence-specific manner to induce the degradation. In addition, the non-degradable control, NMeLCL-ER(dec), did not induce ERa degradation, suggesting that the ability to bind IAPs is critical for Figure 4. Inhibition of the estrogen-dependent tran- the degradation activity. Studies using UPS inhibitors suggested scriptional activity of ERa by LCL-ER(dec). MCF-7 cells that LCL-ER(dec) degraded the ERa via the UPS. Recently, were transiently transfected with a luciferase reporter plas- TF-PROTACs16 (O′PROTACs17) targeting TFs using double- mid containing three tandem copies of the ERE and control Renilla luciferase plasmid-SV40. After 24 h, the cells were stranded decoy ligands have been reported, indicating that decoy-type PROTACs are also useful as a new type of POI in- further transfected with 10 µM concentrations of the indi- ducer. In the future, we aim to apply the methods developed in cated decoys in the presence of 0.3 nM b-estradiol for 24 h. this study to the design of degradation inducers for transcrip- The ERa-dependent transcriptional activity was evaluated tion-related factors that are difficult to target. by a luciferase assay, and the relative luciferase activity was normalized by designating the activity of the non-treated control (column 1) as 100%. The data represent the mean ± S.D. (n = 4). P values were determined using the unpaired ASSOCIATED CONTENT two-tailed Student’s t-test. N.S., not significant; ** P < proteasome0.05; *** P inhibitor < 0.001. MG132, and the ubiquitin-activating in- Supporting Information hibitor MLN7243 (Figure 3C), suggesting that LCL-ER(dec) The Supporting Information is available free of charge on the ACS induced ubiquitin-proteasome system (UPS)-dependent degra- Publications website at DOI: . dation of the ERa. Unlike LCL-ER(dec), ER(dec) did not induce degradation of the ERa protein (Figure 3D), indicating The synthetic procedures for all compounds listed in this manu- that conjugation of the LCL161 ligand is important for the deg- script and the protocols for the in vitro assays (binding and protein radation. In addition, the non-degradable control NMeLCL- degradation assays) (PDF). ER(dec), which contains an N-methylated analog of the LCL161 ligand, did not induce ERa degradation (Figure 3E), suggesting that the ability to bind IAPs is critical for the degra- AUTHOR INFORMATION dation activity. To examine whether target recognition by the Corresponding Author decoy ligands was DNA-sequence dependent, we synthesized * Tel: +81-44-270-6578, Fax: +81-44-270-6578, E-mail: LCL161 ligand-decoy chimeras using several different decoys [email protected] (Y. Demizu). Tel: +81-44-270-6537, E-mail: composed of scrambled DNA sequences as the target ligands. [email protected] (N. Ohoka). These decoy chimeras did not show significant ERa- degradation activity (Figure 3F), which correlated with their Author Contributions weak binding affinity to the ERa (Table 1). These results sug- Miyako Naganuma, N.O., G.T., and H.T. performed the experi- gested that decoy oligonucleotides can be used in the develop- ments and analyzed results. Miyako Naganuma, N.O., T.I., Mik- ment of chimeric molecules that induce DNA sequence-de- ihiko Naito, and Y.D. designed the research and wrote the paper. pendent degradation of target proteins. All authors discussed the results and commented on the manuscript. Because the ERa plays an essential role in estrogen sig- ORCID naling, we examined the effect of LCL-ER(dec) on the estro- Miyako Naganuma : 0000-0003-0011-6268 gen-dependent transcriptional activity of the ERa. In a lucifer- Nobumichi Ohoka : 0000-0002-0533-0610 ase assay using an ERE reporter, LCL-ER(dec) inhibited ERa- dependent transcriptional activation by b-estradiol more effec- Takao Inoue : 0000-0002-6507-9480 tively than ER(dec), which was in good agreement with the Mikihiko Naito : 0000-0003-0451-1337 ERa degradation activity (Figure 4). A mixture of the decoy Yosuke Demizu : 0000-0001-7521-4861 ligand ER(dec) and LCL-PEG3-N3 did not effectively de- crease the transcription, indicating that the linking of the two Funding Sources ligands, ER(dec) and LCL-PEG3-N3, is crucial for effective This study was supported in part by grants from AMED under grant inhibition. These results suggested that degradation of the ERa numbers JP21mk0101197 (to Y.D. and N.O.) and JP21ak0101073 by LCL-ER(dec) leads to effective suppression of estrogen sig- (to N.O., T.I. and Y.D.), 21mk0101187 (to N.O. and T.I.) and naling. JP21am0401003 (to T.I.), the Japan Society for the Promotion of

Science (KAKENHI, grants JP21K05320 to Y.D.; JP18H05502 to (8) Morishita, R.; Higaki, J.; Tomita, N.; Ogihara, T., Applica- Mikihiko Naito and Y.D.; JP18K06567 and JP21K06490 to N.O.; tion of transcription factor "decoy" strategy as means of gene JSPS Fellows 2121J23036 to Miyako Naganuma), Terumo Foun- therapy and study of gene expression in cardiovascular dis- dation for Life Sciences and Arts (to Y.D.), Takeda Science Foun- ease. Circ. Res. 1998, 82 (10), 1023-1028. dation (to Y.D.), the Naito Foundation (to Y.D.), the Sumitomo (9) Pawlak, M.; Lefebvre, P.; Staels, B., General Molecular Bi- Foundation (to Y.D.), the Novartis Foundation (Japan) for the Pro- ology and Architecture of Nuclear Receptors. Curr. Top. Med. motion of Science (to Y.D.), Japan Foundation of Applied Enzy- Chem. 2012, 12 (6), 486-504. mology (to Y.D.), Kobayashi Foundation for Cancer Research (to (10) Kumar, V.; Chambon, P., THE ESTROGEN-RECEPTOR Y.D.) and Foundation for Promotion of Cancer Research in Japan BINDS TIGHTLY TO ITS RESPONSIVE ELEMENT AS A (to Y.D.). LIGAND-INDUCED HOMODIMER. Cell 1988, 55 (1), 145-156. (11) Ohoka, N.; Okuhira, K.; Ito, M.; Nagai, K.; Shibata, N.; Hat- ACKNOWLEDGMENT tori, T.; Ujikawa, O.; Shimokawa, K.; Sano, O.; Koyama, R.; We thank Edanz (https://www.jp.edanz.com/ac) for editing a draft Fujita, H.; Teratani, M.; Matsumoto, H.; Imaeda, Y.; Nara, of this manuscript. H.; Cho, N.; Naito, M., In Vivo Knockdown of Pathogenic Proteins via Specific and Nongenetic Inhibitor of Apoptosis ABBREVIATIONS Protein (IAP)-dependent Protein Erasers (SNIPERs). J. Biol. Chem. 2017, 292 (11), 4556-4570. PROTACs, proteolysis targeting chimeras; IAP, inhibi- (12) Ohoka, N.; Morita, Y.; Nagai, K.; Shimokawa, K.; Ujikawa, tor of apoptosis protein; SNIPERs, specific and non-genetic IAP- O.; Fujimori, I.; Ito, M.; Hayase, Y.; Okuhira, K.; Shibata, dependent protein erasers; UPS, ubiquitin proteasome system; TF, N.; Hattori, T.; Sameshima, T.; Sano, O.; Koyama, R.; transcription factor; ERa, estrogen receptor a; POI, protein of in- Imaeda, Y.; Nara, H.; Cho, N.; Naito, M., Derivatization of terest; VHL, von Hippel-Lindau protein; NOTCH1, neurogenic lo- inhibitor of apoptosis protein (IAP) ligands yields improved cus notch homolog protein 1; 4-OHT, 4-hydroxytamoxifen; ERE, inducers of estrogen receptor α degradation. J. Biol. Chem. estrogen-responsive element; FAM, fluorescein; CD, circular di- 2018, 293 (18), 6776-6790. chroism; Tm, melting temperature. (13) Kleinhitpass, L.; Schorpp, M.; Wagner, U.; Ryffel, G. U., AN ESTROGEN-RESPONSIVE ELEMENT DERIVED FROM REFERENCES THE 5' FLANKING REGION OF THE XENOPUS (1) Ferguson, F. M.; Gray, N. S., Kinase inhibitors: the road VITELLOGENIN A2 GENE FUNCTIONS IN ahead. Nat. Rev. Drug Discov. 2018, 17 (5), 353-376. TRANSFECTED HUMAN-CELLS. Cell 1986, 46 (7), 1053- (2) Nelson, A. L.; Dhimolea, E.; Reichert, J. M., Development 1061. trends for human monoclonal antibody therapeutics. Nat. Rev. (14) Schwabe, J. W. R.; Chapman, L.; Finch, J. T.; Rhodes, D., Drug Discov. 2010, 9 (10), 767-774. THE CRYSTAL-STRUCTURE OF THE ESTROGEN- (3) Burslem, G. M.; Crews, C. M., Small-Molecule Modulation RECEPTOR DNA-BINDING DOMAIN BOUND TO DNA of Protein Homeostasis. Chem. Rev. 2017, 117 (17), 11269- - HOW RECEPTORS DISCRIMINATE BETWEEN THEIR 11301. RESPONSE ELEMENTS. Cell 1993, 75 (3), 567-578. (4) Itoh, Y.; Ishikawa, M.; Naito, M.; Hashimoto, Y., Protein (15) Zengerle, M.; Chan, K. H.; Ciulli, A., Selective Small Mole- Knockdown Using Methyl Bestatin-Ligand Hybrid Mole- cule Induced Degradation of the BET Bromodomain Protein cules: Design and Synthesis of Inducers of Ubiquitination- BRD4. ACS Chem. Biol. 2015, 10 (8), 1770-1777. Mediated Degradation of Cellular Retinoic Acid-Binding (16) Zhang, F. Q.; Wu, Z. W.; Chen, P.; Zhang, J.; Wang, T.; Zhou, Proteins. J. Am. Chem. Soc. 2010, 132 (16), 5820-5826. J. P.; Zhang, H. B., Discovery of a new class of PROTAC (5) Yokoo, H.; Ohoka, N.; Naito, M.; Demizu, Y., Design and BRD4 degraders based on a dihydroquinazolinone derivative synthesis of peptide-based chimeric molecules to induce deg- and lenalidomide/pomalidomide. Bioorg. Med. Chem. 2020, radation of the estrogen and androgen receptors. Bioorg. Med. 28 (1). Chem. 2020, 28 (15), 115595. (17) Liu, J.; Chen, H.; Kaniskan, H. U.; Xie, L.; Chen, X.; Jin, J.; (6) Ohoka, N.; Misawa, T.; Kurihara, M.; Demizu, Y.; Naito, M., Wei, W. Y., TF-PROTACs Enable Targeted Degradation of Development of a peptide-based inducer of protein degrada- Transcription Factors. J. Am. Chem. Soc. 2021, 143 (23), tion targeting NOTCH1. Bioorg. Med. Chem. Lett. 2017, 27 8902-8910. (22), 4985-4988. (18) Shao, J.; Yan, Y.; Ding, D.; Wang, D.; He, Y.; Pan, Y.; Yan, (7) Keefe, A. D.; Pai, S.; Ellington, A., Aptamers as therapeutics. W.; Kharbanda, A.; Li, H. Y.; Huang, H., Destruction of Nat. Rev. Drug Discov. 2010, 9 (7), 537-550. DNA-Binding Proteins by Programmable Oligonucleotide PROTAC (O'PROTAC): Effective Targeting of LEF1 and ERG. Adv. Sci. 2021, e2102555.

For Table of Contents Graphic Use Only

Development of chimeric molecules that degrade the estrogen receptor using decoy oligonucleotide ligands

Miyako Naganuma, Nobumichi Ohoka, Genichiro Tsuji, Haruna Tsujimura, Kenji Matsuno, Takao Inoue, Mikihiko Naito, and Yosuke Demizu

N N N

N O N S O N O Me H Me N Me

UPS-mediated degradation

Lay Summary

Protein degradation technology using hybrid small molecules (known as PROTACs and SNIPERs) is an emerging drug development strategy for use in drug discovery and biological research. In this study, a decoy ligand-based estrogen receptor a (ERa) degrader LCL-ER(dec) was designed and synthesized and the binding affinity and degradation activity toward the ERa were evaluated. LCL-ER(dec) was revealed to selectively degrade the ERa via the ubiquitin-proteasome system. Because we were able to show that decoys were able to be used as the ligands for protein degraders, we expect that this concept will lead to the expansion of the number of proteins that can be targeted.